During the course of an immune response, activated B cells either differentiate into antibody- forming cells (AFC) or seed germinal centers. AFC constitute the bulk of the primary immune B-cell response and are characterized by IgM antibodies early on, with some switching to downstream isotypes such as IgG late in a persistent infection. Germinal center B-cells, on the other hand, contribute little to the primary response: their main role is to undergo a process of hypermutation aimed at changing the specificity of the immunoglobulin receptor. Hypermutation is coupled to a cellular selection mechanism for high-affinity variants and this coupling leads to the generation of high-affinity memory B cells. Memory B cells contribute greatly to the secondary immune response and are, along with the memory T cell response, the reason why we are less likely to experience major symptoms of infection upon re-exposure to a particular antigen. The molecular basis of the immunoglobulin hypermutation mechanism is unknown and is one of the preoccupations of this laboratory. A novel cytosine deaminase termed AID was recently found to play a pivotal role in immunoglobulin hypermutation. AID appears to work by deaminating cytosines in DNA, leading to uracil in the DNA. Uracil in DNA can be dealt with by one of the uracil DNA glycosylases, or it is read as thymine during replication, leading to G:C to A:T transitions. Uniquely in immunoglobulin hypermutation, the evidence suggests that the presence of the G:U mismatch or the G:Ap site (Ap = abasic) leads to the recruitment of translesion synthesis DNA polymerases Eta, Iota and Zeta, leading to the generation of mutations at all bases, not just the G:C targeting expected from cytosine deamination on both strands. The repeated introduction of uracil into the DNA of immunoglobulin genes by AID and the resulting recruitment of the error-prone polymerases leads to a mutation frequency that is more than 1 million fold over background. We have recently found evidence that AID targets the immunoglobulin hotspot, DGYW, and demonstrated that AID is regulated by a nuclear export mechanism. We also demonstrated that AID does not intrinsically bind DNA but that it probably requires interaction with a protein partner for efficient localization and DNA binding. We are currently investigating the identity of this partner, as well as continuing to examine the role of error-prone DNA polymerases in immunoglobulin hypermutation by the generation of various genetically altered mouse strains.